PROJECT SUMMARY/ABSTRACT Derivation of stable and chimera competent embryonic stem cells (ESCs) from several rodent species has revolutionized modern medicine. Germline transmission of genetically modified ESCs has become a power technology for modeling human diseases in live rodents. In addition, transplantation of rodent ESC-derived cells, tissues and organs provides a testbed for repairing or replacing damaged tissues and organs in the human body, a holy grail for regenerative medicine. However, there are considerable differences between rodents and humans in terms of body size, lifespan and physiology, which limit the degree to which insights derived from the rodent models can be applied to humans. Large animal models often have an enhanced ability to predict clinical efficacy relative to rodents. Despite numerous attempts, however, bona fide ESC analogous to those described for rodents are still not available in any large animal species. Recently we developed a new method that enabled, for the first time, the derivation of stable and chimera competent ESCs from a large domesticated mammal, the horse. The objective of this proposal is to apply the newly developed method for the derivation of stable and chimera competent ESCs from one of the most popular large animal models in biomedical research, the pig. Chimera competent pig ESCs will not only facilitate the production of transgenic pig models of human diseases but also provide an unprecedented opportunity to test the efficacy and safety of blastocyst complementation in a large animal model for in vivo generation of tissues and organs for transplantation, a necessary step prior to human applications.